Cytometry Lab

Common Flow Cytometry Fluorochrome Combinations

Standard or reasonably common fluorochrome combinations used in WEHI instruments:

• FACStar+
• FACSVantageSEDiVa
• MoFlo
• Aria
• LSR I
• LSR II
• FACSCalibur
• FACScan

FACStar+ Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3*/FL5**	PI, 7AAD, PE.Tx , PE.Cy5, PerCp.Cy5.5, PE.Cy7
		FL6**	PE.Cy7***
	515nm***	FL2	TRITC
Laser 2	605nm	FL3*	APC, Cy5, Alexa630, Alexa647, Alexa660,
		FL4	Tx.Red, Alexa594
	550-650nm	FL3*, FL4	dye laser tuned for special fluorochrome excitation

• * FL3 may accept laser1 or laser2-excited signals
• ** When set for 8-parameter analysis
• *** Allow 15 minutes setup time
• PE.Cy5 should not be combined with any FL3/laser 2 fluorochrome
• PE.Tx should not be combined with any FL4 fluorochrome

DiVa Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3	7AAD, PE.Cy5, PerCp.Cy5.5, PE.Cy7
		FL6	PI, PE.Tx, PE.Alexa610
	515nm*	FL2	TRITC
Laser 2	605nm	FL4	Tx.Red, Alexa594, APC.Cy7*, Alexa680*
		FL5	APC, Cy5, Alexa630, Alexa647, Alexa660
	550-650nm	FL4/FL5	dye laser tuned for special fluorochrome excitation
Laser 3	351-364nm	FL7	Hoechst, Alexa350, Cascade Blue, AMCA, Indo-1, DAPI, Qdot605*
		FL8	Indo-1, CascadeYellow, FluoroGold, HoechstRed (SP cells), Qdot655*

• * Allow 15 minutes setup time
• PE.Cy5 should not be combined with any FL5 fluorochrome
• PE.Tx should not be combined with any FL4 fluorochrome

MoFlo Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3	PI, PE.Tx, PE.Alexa610
		FL4	PE.Cy5, PerCp.Cy5.5, PE.Cy7, 7AAD
Laser 2	632nm	FL5	APC, Cy5, Alexa630, Alexa647, Alexa660
		FL6	APC.Cy7
Laser 3	351-364nm	FL7	Hoechst, Alexa350, Cascade Blue, AMCA, Indo-1, DAPI, Qdot605*
		FL8	Indo-1, CascadeYellow, FluoroGold, HoechstRed (SP cells), Qdot655*
	405nm**	FL7	Alexa405

• * Allow 15 minutes setup time
• ** Allow 30 minutes setup time
• PE.Cy5 should not be combined with any FL5 fluorochrome

Aria Laser and Fluorochrome Options:

Laser 1	488nm	E	FITC, GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		D	PE, Cy3, RFP
		C	PI, PE.Tx, PE.Alexa610, 7AAD, PE.Cy5
		B	PerCp.Cy5.5
		A	PE.Cy7
Laser 2	405nm	E	Alexa 405, Pacific Blue
		D	525nm
		C	CascadeYellow
		B	585nm
		A	QDot655
Laser 3	375nm	C	Hoechst Blue, Alexa350, Cascade Blue, AMCA, DAPI
		B	FluoroGold
		A	Hoechst Red (SP cells)
Laser 4	635nm	B	APC, Cy5, Alexa630, Alexa647
		A	APC.Cy7, Alexa 680, Alexa 700

LSR1 Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3	PI, 7AAD, PE.Tx , PE.Cy5, PerCp.Cy5.5, PE.Cy7
Laser 2	325nm	FL4	Indo-1, CascadeYellow, FluoroGold
		FL5	Hoechst, Alexa350, Cascade Blue, AMCA, Indo-1, DAPI
Laser 3	632nm	FL6/SSC-W	APC, Cy5, Alexa630, Alexa647, Alexa660
		FL3*	Alexa680, APC.Cy7

• * Special setup for laser 3 FL3 excitation
• PE.Cy5 should not be combined with any FL6 fluorochrome

LSR II Laser and Fluorochrome Options:

Laser 1	488nm	E	FITC, GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		D	PE, Cy3, RFP
		C	PI, PE.Tx, PE.Alexa610, 7AAD
		B	PerCp.Cy5.5, PE.Cy5
		A	PE.Cy7
Laser 2	355nm	D	Hoechst Blue, INDO-1 blue, Alexa 350
		C	Indo-1 green, Cascade Yellow, FluoroGold
		B	QDot605
		A	QDot655, Hoechst Red (SP cells)
Laser 3	635nm	B	APC, Cy5, Alexa630, Alexa647
		A	APC.Cy7, Alexa 680, Alexa 700

FACSCalibur Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3	PI, 7AAD, PE.Tx , PE.Cy5, PerCp.Cy5.5, PE.Cy7
Laser 2	635nm	FL4	APC, Cy5, Alexa630, Alexa647, Alexa660

• PE.Cy5 should not be combined with any FL4 fluorochrome

FACSCan 1 and 2 Laser and Fluorochrome Options:

Laser 1	488nm	FL1	FITC , GFP, CFSE, 5'6Carboxyfluorescein, Alexa488, YFP
		FL2	PE, Cy3, RFP
		FL3	PI, 7AAD, PE.Tx , PE.Cy5, PerCp.Cy5.5, PE.Cy7

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